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. Author manuscript; available in PMC: 2013 Mar 1.
Published in final edited form as: Dev Dyn. 2012 Jan 30;241(3):505–521. doi: 10.1002/dvdy.23735

Figure 6.

Figure 6

TEG-1 directly interacts with UAF-1 in C. elegans extracts, and this interaction is conserved in humans. (A) Reciprocal co-immunoprecipitations (IPs) were performed using UAF-1 specific antibodies (left) and TEG-1 specific antibodies (right), followed by SDS electrophoresis and western blotting. In the UAF-1 IP, TEG-1 specific antibodies detect a band of the correct size, whereas an IP using non-specific pre-immunized rabbit serum, did not result in detection of TEG-1. In the TEG-1 IP, a band of the correct size was detected with UAF-1 specific antibodies, but not when pre-immune serum was used to perform the IP. (B) E. coli expressed GST-TEG-1 and 6X-His-UAF-1 co-IP. 6X-His-UAF-1 was expressed in E. coli cells with GST, GST-TEG-1, or GST-TEG-1 with the GYF domain removed (GST-TEG-1(ΔGYF); Figure 3B). When UAF-1 specific antibodies were used to IP, only GST-TEG-1(full length) was co-IP’d (top). Likewise, when GST specific antibodies where used to IP, only the strain contain GST-TEG-1(full length) was able to co-IP UAF-1, as detected by UAF-1 specific antibodies. (C) Co-IP of U2AF65 and CD2BP2 from HeLa extracts. GFP or GFP-CD2BP2 were transiently expressed in HeLa cells and proteins were IP’d using anti-GFP specific antibodies (left) or anti-U2AF65 specific antibodies (right). U2AF65 and U2AF35 were detected in the anti-GFP IP on a western blot, while the negative control, Hsp90 heat shock protein, was not co-IP’d. Likewise, GFP-CD2BP2 was co-IP’d with U2AF65, while GFP and Hsp90 were not (right). U2AF35 was co-IP’d with U2AF65, whether using lysate from GFP expressing cells, or GFP-CD2BP2 expressing cells. (D) E. coli expressed GST-CD2BP2 and U2AF65 co-IP. U2AF65 was expressed in E. coli cells with GST, GST-CD2BP2(full length), or GST-CD2BP2(ΔGYF). Only GST-CD2BP2(full length) was co-IP’d with U2AF65. Likewise, when IP was performed using anti-GST antibodies, U2AF65 was only detected when lysate from cells expressing GST-CD2BP2(full length) was used; U2AF65 could not be detected when lysate from cells expressing GST or GST-CD2BP2(ΔGYF) was used for the IP.